Open AccessA protein factor of rat liver mitochondrial matrix involved in flavinylation of dimethylglycine dehydrogenase
Author(s) -
Brizio Carmen,
Otto Annegret,
Brandsch Roderich,
Passarella Salvatore,
Barile Maria
Publication year - 2000
Publication title -
european journal of biochemistry
Language(s) - English
Resource type - Journals
eISSN - 1432-1033
pISSN - 0014-2956
DOI - 10.1046/j.1432-1327.2000.01464.x
Subject(s) - mitochondrial matrix , biochemistry , mitochondrion , hsp60 , inner mitochondrial membrane , chemistry , reticulocyte , biology , microbiology and biotechnology , cytosol , enzyme , heat shock protein , hsp70 , rna , gene
The involvement of rat liver mitochondria in the flavinylation of the mitochondrial matrix flavoenzyme dimethylglycine dehydrogenase (Me 2 GlyDH) has been investigated. Me 2 GlyDH was synthesized as an apoenzyme in the rabbit reticulocyte lysate (RL) transcription/translation system and its flavinylation was monitored by virtue of the trypsin resistance of the holoenzyme. The rate of holoenzyme formation in the presence of FAD was stimulated with increasing efficiency by the addition of solubilized mitoplasts, mitochondrial matrix and DEAE‐purified matrix fraction. Apo‐Me 2 GlyDH was also converted into holoenzyme when the solubilized mitoplasts were supplemented with FMN and ATP. This observation is consistent with the existence of a mitochondrial FAD synthetase generating the FAD needed for holoenzyme formation from its precursors. Holoenzyme formation in the presence of FAD increased linearly with the concentration of matrix protein in the assay, and depended on the amount of externally added Me 2 GlyDH with saturation characteristics. These findings suggest the presence of a protein factor in the mitochondrial matrix which stimulates Me 2 GlyDH flavinylation. This factor was different from both mitochondrial heat shock protein (Hsp)70, as shown by immunodepletion experiments, and mitochondrial Hsp60, as demonstrated by the capability of a DEAE‐purified matrix fraction devoid of Hsp60 to accelerate flavinylation of both RL translated and purified Me 2 GlyDH.